Human monoclonal antibody against Hepatitis B virus surface antigen (HBVSAG)

ABSTRACT

Disclosed is a hybridoma cell line which produces human antibodies capable of binding to the hepatitis B virus surface antigen (HBVsAg), as well as antibodies produced by the cell line. Also disclosed are various uses of said antibodies in the prevention and treatment of HBV infection. Peripheral blood lymphocytes obtained from human donors having a high titer of anti HBVsAg antibodies are activated in vitro with pokeweed mitogen and then fused with heteromyeloma cells to generate hybridomas secreting human antibodies having a high affinity and specificity to HBVsAg.

FIELD OF THE INVENTION

The present invention concerns a hybridoma cell line producing humanantibodies capable of binding to the hepatitis B virus surface antigen,antibodies produced by the cell lines, and various uses thereof.

BACKGROUND OF THE INVENTION

Hepatitis B virus (HBV) infection is a major worldwide health problem.Approximately 5% of the world population is infected by HBV andchronically infected patients carry a high risk of developing cirrhosisand hepatocellular carcinoma. (Progressive Hepatitis Research: HepatitisB virus (HBV), Hepatitis C virus (HCV) and Hepatitis Delta virus (HDV)Ed. O. Crivelli, Sorina Biomedica, 1991).

The immune response to HBV-encoded antigens includes both a cellularimmune response which is active in the elimination of HBV infectedcells, as well as a humoral antibody response to viral envelope antigenswhich contributes to the clearance of circulating virus particles. Thedominant cause of viral persistence during HBV infection is thedevelopment of a weak antiviral immune response.

Recombinant HBV vaccines provide a safe and effective means for activeimmunization against HBV, however, they do not always induce asufficient and rapid antibody response.

Interferon-α has been used in the therapy of Hepatitis B infectionshowing an efficacy of only 30-40% in highly selected patients.

In addition, passive immunization with human polyclonal anti Hepatitis Bantisera has been shown to be effective in delaying and even preventingrecurrent HBV infection (Wright, T. L. and Lau, J. Y. N. The Lancet342:1340-1344, (1993)). Such human polyclonal antisera are prepared frompooled plasma of immunized donors. These preparations are very expensiveand available in relatively small amounts. Furthermore, pooled plasmamay contain contaminated blood samples and thus treatment with suchantisera increases the patient's risk to contract other viral infectionssuch as hepatitis C or HIV.

An alternative approach for the treatment of HBV infections concerns theuse of monoclonal antibodies (MoAb).

PCT patent application PCT/NL94/00102 discloses human monoclonalantibodies directed against Hepatitis B surface antigen which aresecreted by the hybridoma cell lines Mab 4-7B and Mab 9H9. Themonoclonal antibody secreted by the cell line Mab 4-7B recognizes alinear epitope of HBVsAg and is different from the Mab 9H9 monoclonalantibody which recognizes a conformational epitope. The antibodies areclaimed for simultaneous use in the treatment of chronic Hepatitis Binfections.

PCT patent application PCT/US92/09749 discloses human monoclonalantibodies against HBVsAg which are secreted by the hybridoma cell linesPE1-1, ZM1-1, ZM1-2, MD3-4 and LO3-3. The antibodies bind to differentHBV epitopes and are used for reducing the level of circulating HBVsAg.

Japanese Patent Application JP 93066104 discloses a hybridoma of a humanlymphocyte cell strain TAW-925 and a human lymphocyte transformed byEpstein-Barr virus. The hybridoma produces a human monoclonal antibodyagainst HBVsAg.

U.S. patent application Ser. No. 4,883,752 discloses preparation ofhuman-derived monoclonal antibody to HBVsAg, by administration of HBVsAgvaccine to humans, recovering their lymphocytes, stimulating thelymphocytes in vitro by a non specific stimulator, fusing said cellswith a myeloma cell, and selecting for hybridomas with secrete andHBVsAg antibodies.

Ichimori et al., Biochem. and Biophysic. Research Communications129(1):26-33, 1985 discloses a hybridoma secreting human anti HBVsAgmonoclonal antibodies which recognize the a-determinant of HBVsAg.Later, Ichimori, et al., supra 142(3):805-812, 1987 disclosed anotherhybridoma which stably secretes human monoclonal antibody against HBsAg.

SUMMARY OF THE INVENTION

In accordance with the present invention, a hybridoma cell line isprovided which secretes human antibodies capable of binding to theHepatitis B surface antigen (HBVsAg).

In accordance with the invention, peripheral blood lymphocytes (PBL)were obtained from human individuals having a high titer of anti HBVsAgantibodies. Such individuals may either have been previously infectedwith HBV, actively immunized with HBV antigens or spontaneously showinga high level of such antibodies. A most preferred human donor is anindividual which tested negative for the presence of HBV but shows ahigh titer of antibodies against HBVsAg. PBLs from the human donor maybe obtained either by whole blood donation or by leukophoresis.

The human PBLs are then activated in vitro by their incubation withpokeweed mitogen (PWM). After activation the PBLs are fused in vitropreferably with a human-mouse fusion partner such as a heteromyeloma bytechniques well known in the art (e.g. Kohler & Milstein, Nature,256:495-497, 1975). The generated hybridoma cell lines are eithercultured in vitro in a suitable medium wherein the desired monoclonalantibody is recovered from the supernatant or, alternatively thehybridoma cell lines may be injected intraperitoneally into mice and theantibodies harvested from the malignant ascitis or serum of these mice.The supernatant of the hybridoma cell lines are first screened forproduction of human IgG antibodies by any of the methods known in theart such as enzyme linked immunosorbent assay (ELISA) or radioimmunoassay (RIA). Hybridomas testing positive for human IgG are then furtherscreened for production of anti HBVsAg antibodies by their capability tobind to HBVsAg.

In accordance with the preferred embodiment of the present invention, ahybridoma cell line designated herein as “17.1.41” which was depositedon May 22, 1996, at the European Collection of Cell Cultures (ECACC,CAMR, Salisbury, Wiltshire, SP40JG, U.K.) under the Accession No.96052169 is provided. Anti HBVsAg human monoclonal antibodies secretedby the above hybridoma cell line designated herein as “Ab17.1.41” aswell as fragments thereof retaining the antigen binding characteristicsof the antibodies and antibodies capable of binding to the antigenicepitope bound by Ab17.1.41 are also provided. Such fragments may be, forexample, Fab or F(ab)₂ fragments obtained by digestion of the wholeantibody with various enzymes as known and described extensively in theart. The antigenic characteristics of an antibody are determined bytesting the binding of an antibody to a certain antigenic determinantusing standard assays such as RIA, ELISA or FACS analysis.

The antibodies of the invention have a relatively high affinity toHBVsAg being in the range of about 10⁻⁹M to about 10⁻¹⁰M as determinedby a competitive ELISA assay.

The antigen bound by the antibodies defined above also constitutes anaspect of the invention.

Further aspects of the present invention are various diagnosticprophylactic and therapeutic uses of the Ab 17.1.41 monoclonalantibodies and the Ag bound by these antibodies. In accordance with thisaspect of the invention, pharmaceutical compositions comprising theAb17.1.41 antibodies may be used for the treatment of chronic HepatitisB patients by administering to such a patient a therapeuticallyeffective amount of the antibodies or fragments thereof capable ofbinding to the HBVsAg being an amount effective in alleviating thesymptoms of the HBV infection or reducing the number of circulatingviral particles in an individual.

In addition to the antibodies of the invention the pharmaceuticalcompositions may optionally also comprise a carrier selected from any ofthe carriers known in the art. One example of such a carrier is aliposome. The pharmaceutical compositions of the invention may alsocomprise various diluents and adjuvants known per se.

The compositions of the invention may be administered by a variety ofadministration modes including parenterally, orally etc. Compositionscomprising the antibodies of the invention, as described above, may beadministered in combination with other anti viral agents. Such agentsmay include, as a non limiting example: Interferons, anti HB monoclonalantibodies, anti HB polyclonal antibodies, nucleoside analogs, andinhibitors of DNA polymerase. In the case of such a combination therapythe antibodies may be given simultaneously with the anti viral agent orsequentially either before or after treatment with the anti viral agent.

Such pharmaceutical compositions may also be used, for example, forimmunization of new born babies against HBV infections or forimmunization cf of liver transplantation patients to eliminate possiblerecurrent HBV infections in such patients.

By a further embodiment, the antibodies of the invention may also beused in a method for the diagnosis of HBV infections in an individual byobtaining a body fluid sample from the tested individual which may be ablood sample, a lymph sample or any other body fluid sample andcontacting the body fluid sample with a human anti HBVsAg antibody ofthe invention under conditions enabling the formation ofantibody-antigen complexes. The level of such complexes is thendetermined by methods known in the art, a level significantly higherthan that formed in a control sample indicating an HV infection in thetested individual. In the same manner, the specific antigen bound by theantibodies of the invention may also be used for diagnosis of HBinfection in an individual by contacting a body fluid sample from thetested individual with the antigen as described above and determiningthe formation of antigen Ab in the sample.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a photograph showing Hepatitis B infected liver sectionsstained with the anti HBVs antibodies of the invention. All sectionswere stained with a “secondary” antibody, i.e. goat anti human or antimouse Ig conjugated to biotin.

-   A—negative control. No first antibody.-   B—positive control. First antibody—mouse anti HB antibody and a    secondary anti-mouse Ig.-   C—staining with anti HBVsAg Ab 17.1.41.

Reference will now be made to the following Examples which are providedby way of illustration and are not intended to be limiting to thepresent invention.

FIG. 2 is a schematic representation of the binding of Ab17.1.41 to aset of well characterized HBsAg types. The y axis represents opticaldensity units. The x axis represents different HBsAg types.

FIG. 3 is a graphic representation of hepatitis B viremia score, asdefined in example 3. Each dot in the graph represents one animal.

FIG. 4 is a graphic representation of the percentage of HBV infectedanimals at days 18 and 25 in the untreated group and Ab17.1.41 treatedgroup (in the treatment model).

FIG. 5 is a graphic representation of the percentage of HBV infectedanimals at days 10 and 17 in the untreated group and Ab17.1.41 treatedgroup (in the combined prophylaxis/inhibition model).

FIG. 6 is a graphic representation of the percentage of HBV infectedanimals at days 11 and 18 in the untreated group and Ab 17.1.41 treatedgroup (in the combined inhibition/treatment model).

FIG. 7 is a graphic representation of the percentage of HBV infectedanimals at days 21 and 27 in the untreated group (control), the grouptreated with an anti viral drug, the group treated with Ab17.1.41 andthe group treated with both the anti viral drug and Ab17.1.41(Mab+Drug).

FIG. 8 Nucleic acid sequence (SEQ ID NO:1) and corresponding amino acidsequence (SEQ ID NO:2) of the light chain of the variable domain ofAb17.1.4 1.

FIG. 9 Nucleic acid sequence (SEQ ID NO:3) and corresponding amino acidsequence (SEQ ID NO:4) of the heavy chain of the variable domain of Ab17.1.41.

EXAMPLES

Materials and Methods

In vitro Activation:

Peripheral blood lymphocytes (PBL) were obtained after informed consentby leukophoresis from donors positive for HBs antibodies and negativefor HBV. PBLs were washed twice, counted and resuspended in PBS to thedesired cell concentration. PBL were separated from granulocytes anderythrocytes on a Ficoll-hypaque gradient (UNI-SEP maxi; Eldan Tech.,Jerusalem, Israel) and subsequently stimulated for 3-4 days withpokeweed mitogen (PWM; Gibco BRL, Life Technologies Inc., Grand Island,N.Y.) diluted 1:100 and with Antigen at 200 ng/ml in RPMI-1640 mediumwith 10% (v/v) fetal calf serum (FCS) supplemented with 10 U/mlpenicillin, 10 μg/ml streptomycin, 2 mM L-glutamine, 1 mM sodiumpyruvate, 1% (v/v) non-essential amino acids (Biological Industries,Belt Haemek, Israel) and 10⁻⁴ M 2-mercaptoethanol (Sigma, St. Louis)(Complete Medium).

Cell Fusion:

Cells were mixed with the human-mouse heteromyeloma HMMA2.1 1TG/0(Posner et al.) at 3:1 ratio. Fusion was performed with 50% (w/v) PEG1500 (Boehringer Manheim GmbH) in a standard procedure. Fused cells wereseeded at a concentration of 30000 cells/well in 96-well U-bottommicrotiter plates (Nunc, Denmark) in complete medium containingHAT-supplement (1×) (Biological Industries, Beit Haemek, Israel). Cellswere fed with fresh HAT-medium a week latter. Two weeks after fusionsupernatants were harvested for ELISA and medium was replaced with freshHT-medium.

Hybridoma cultures secreting specific anti-HBs Ig were cloned at 0.5cell/well in 96-well U-bottom microtiter plates.

Determination of Human Immunoglobulin:

Sera were tested for antigen specific and total human ig. Total human Igwas quantified by sandwich ELISA using goat F(ab)₂-purified anti-humanIgG+IgM+IgA (Zymed Laboratories, San Francisco, Calif.) as the captureagent and peroxidase-conjugated purified goat anti-human (ZymedLaboratories) as the detection reagent. Human serum of knownimmunoglobulin concentration was used as the standard (Sigma, Rehovot,Israel). Microplates (Nunc, Roskilde, Denmark) pre-coated with thecapture reagent (2.5 ug/ml, 50 ul/well) and blocked with 1% BSA wereincubated overnight at 4C with dilutions of plasma from 1:20000 to1:640000, or the standard from 0.2 to 0.06 ug/ml, then washed 5 timeswith PBS-Tween solution. The detection reagent was added and the plateswere incubated for 1 h at 37C, then washed again 3 times. Freshsubstrate solution (TMB, Sigma) was added and, afterperoxidase-catalyzed color development, the reaction was stopped byaddition of 10% sulfuric acid. Absorbance at 450 nm was quantified on anELISA reader (Dynatech, Port Guernsey, Channel Islands, UK).

Concentration of antigen-specific human antibodies in mice sera wasdetermined by HBsAb EIA kit (ZER, Jerusalem, Israel).

Human antibodies in hybridoma supernatants were determined by overnightincubation of supernatants on goat anti-human IgG+A+M (Zymed) coatedplates, with goat anti-human IgG-peroxidase conjugated as the secondaryreagent.

Antigen-specific antibodies in hybridoma supernatants were determined asabove using Hbs antigen coated plates.

Determination of Human IgG Subclasses:

Human IgG subclasses were determined by sandwich ELISA using goatF(ab)₂-purified anti-human IgG+IgM+IgA (Zymed Laboratories, SanFrancisco, Calif.) coated plates and Hbs antigen coated plates. Mouseanti-human IgG subclasses (Sigma) were used as second antibody andperoxidase-conjugated purified goat anti-human (Zymed Laboratories) asthe detection reagent.Statistic Analysis:

Statistical analysis was performed using the Stat View II program(Abacus Concepts, Inc., Berkeley, Calif.) on a Mackintosh Quadra 605 orMicrosoft Excel 5.0 (Microsoft) on a 486 DX2 PC compatible. Studentt-test, Anova correlation and regression analysis were utilized tocalculate probability (p) and correlation coefficient (r) values.Results are presented as mean ± standard error.

Affinity Constant Measurements:

Determination of affinity constants (K_(D)) of the different anti-HBsantibodies to ad antigen (Chemicon Cat. No. AG 850) in solution wereperformed according to Friguet et al. (Journal of Immunological Methods,77:305-319, 1985). The antigen at various concentrations (3.5×10⁻¹⁰M to1.4×10⁻⁹M) was first incubated in solution with a constant amount ofantibody (3.4×10⁻¹¹M), in 0.1 M sodium phosphate buffer containing 2 mMEDTA and 10 mg/ml BSA, pH 7.8 (medium buffer). After o.n. incubation at20 C the concentration of free antibody was determined by an indirectELISA. A volume of 300 ul of each mixture were transferred and incubatedfor 2 h at 20 C into the wells of a microtitration plate (Nunc)previously coated with Ad (50 μl/well at 1 μg/ml in 0.1 M NaHCO₃ buffer,pH 9.6 for 2 h at 37° C.). After washing with PBS containing 0.04% Tween20, the bound antibodies were detected by adding HRP-F(ab′)₂ Goat antihuman IgG (Zymed) diluted 1:3000 with medium buffer, 50 μl/well 2 h at20° C. The plate was developed with TMB chromogen (Sigma T-3405 tablets)50 μl/well, the reaction stopped with 10% H₂SO₄ 50 μl/well and the plateread in an ELISA reader at 450 nm. The conditions were chosen so thatthe resulting f values (see Friguet et al.) were around 0.1. Theantibody concentration used was deduced from an ELISA calibration doneon the same plate. The affinity constant KD was calculated from therelevant Scatchard plot.

Inhibition Assays:

The inhibition assay was performed in microtiter plates coated with HBsparticles (2 μg/ml in PBS). The plate was blocked with 3% BSA in PBS.Hybridoma supernatants containing anti HBs antibodies were seriallydiluted. 50 μl of each dilution were added to the coated microtiterwells. Subsequently, 50 μl of HBs particles (ad/ay, 0.5 μl/ml in PBS) orPBS alone were added to each well. The plates were incubated overnightat room temperature in a humid chamber and washed 5 times withPBS-Tween. Next, 50 μl of goat anti human IgG conjugated to HRP (diluted1:5000 in PBS) were added to each well. After a 4 hour incubation atroom temperature in a humid chamber the plates were washed 5 times withPBS-Tween, and TMB was added to each well. Results were read using anELISA reader, in a wavelength of 450 nm.

Immunohistostaining:

HBV positive liver fragment was fixed in 4% neutral bufferedformaldehyde for 24 h and then embedded in paraffin using routineprocedures. Section of 4 μm thickness were cut from paraffin blocks andmounted on polylysine-coated slides. After deparaffinization andperoxidase quenching staining was performed using our monoclonal Humananti-HBs Protein A-purified antibodies followed by biotinylated Goatanti-Human IgG (H+L) (Zymed, San Francisco, Calif.) usingHistostain-SPTM kit (Zymed) according to the manufacture'srecommendation. Control slides without using the 1 st Human anti-HBsantibody were stained in parallel.

Sequence analysis:

Total RNA was isolated from 10×10⁶ hybridoma cells with RNAsol B reagent(TEL-TEX, Inc. Friendswood, Tex.). cDNA was prepared from 10 μg of totalRNA with reverse transcriptase and oligo dT (Promega, Madison, Wis.)according to standard procedures. PCR was performed on 1/50 of the RTreaction mixture with V_(H), V_(λ) or V_(K)5′ leader primers and 3′primers corresponding to human constant region. The PCR fragments werecloned into pGEM-T vector (Promega). The inserts were sequenced using anABI 377 sequencing machine. Sequences were analyzed by comparison toGenbank and by alignment to Kabat sequences (Kabat et al. 1991,Sequences of proteins of immunological interest (5^(th) Ed.) U.S. Dept.of Health and Human Services, National Institutes of Health, Bethesda,Md.).

Example 1

Human peripheral blood lymphocytes (PBL) from donors positive for antiHBVs antibodies were obtained and activated in vitro with PWM asdescribed above. The cells were then fused with a human mouseheteromyeloto form hybridoma cell lines. One stable hybridoma clonesecreting specific human anti HBVsAg designated 17.1.41 wascharacterized. The antibodies secreted by the above clone were purifiedon a protein A column as well as on an anti human Ig-agarose column andwere found to be of the IgG1 Vκ type. The affinity constant of theantibodies to HBVsAg was 1.34×10⁻⁹. Specificity was tested bycompetitive inhibition assay using HBV surface antigen of the ad-ay(1:1).

Example 2

The 17.1.41 antibodies were used for staining human liver fragments asdescribed above. As seen in FIG. 1, the 17.1.41 antibodies were able todetect HBV particles present in the infected liver fragments.

The gene encoding the variable region of Ab 17.1.41 was isolated, fullysequenced, and its subgroups and CDRs were determined.

The antibody has a fully human Ig gene sequence as determined byalignment to Genebank sequences and Kabat protein sequences. FIG. 8shows the nucleotide sequence of the cDNA encoding the light chain ofthe variable region of Ab 17.1.41 and its corresponding amino acidsequence (Sequence identification nos. I and 3). FIG. 9 shows thenucleotide sequence of the cDNA encoding the heavy chain of the variableregion of Ab17.1.41 and its corresponding amino acid sequence (Sequenceidentification nos. 2 and 4).

The sequencing data revealed that the variable region of Ab 17.1.41consists of the subgroups V_(H3), J_(H)6, V_(K2) and J_(K2).

HBV genomes are classified into six groups A to F, based on the degreeof similarity in their nucleotide sequences. The genetic variability ofHBV is further reflected in the occurrence of different serotypes ofHBsAg. The common determinant ‘a’ and two pairs of mutually exclusivedeterminants ‘d/y’ and ‘w/r’ enable the distinction of four majorsubtypes of HBsAg: adw, adr, ayw and ayr. Additional determinantsdesignated subdeterminants of w(w1 to w4) have allowed the definition offour serotypes of ayw (ayw1-4) and two serotypes of adw, i.e. adw2 andadw4. Additional subtype variation is added by the q determinant, whichis present on almost all subtypes. Its absence is marked by a ‘q-’ sign.The kind of HBV serotypes recognized by Ab 17.1.41 was examined using aset of 15 different HBsAg types (Norder et al., 1992, Journal of GeneralVirology, 73, 3141; Magnius and Norder, 1995, Intervirology, 38, 24-34).As can be seen in FIG. 2. Ab 17.1.41 has a broad reactivity towards alltested subtypes and genotypes, except for C adw2.

Example 3

The biological activity of Ab 17.1.41 was characterized using thefollowing HBV animal model: a mouse was treated so as to allow thestable engraftment of human liver fragments. The treatment includedintensive irradiation followed by transplantation of scid (severecombined immunodeficient) mice bone marrow. Viral infection of humanliver fragments was performed ex-vivo using HBV positive human serum (EP699 235).

The animal model was used in three different modes representing variouspotential uses of the antibodies: treatment mode, combinedprophylaxis/inhibition mode and combined inhibition/treatment.

-   1. Treatment mode—This model demonstrates the ability to use the    antibody to treat chronic HBV infection. Mice were transplanted with    HBV infected human liver fragments. The mice were treated with Ab    17.1.41 at days 16, and 17 post liver transplantation. HBV DNA was    tested on days 18 and 25. The number of HBV DNA copies (the viral    load) in mouse sera was determined using PCR. We use the term    “viremia score” as a mathematical representation of the viral load.    The viremia score was determined as follows:

Viremia score viral load = HBV DNA copies/ml serum 0 viral load < 5 ×10³ 1 5 × 10³ < viral load < 5 × 10⁴ 2 5 × 10⁴ < viral load < 5 × 10⁵ 3viral load > 5 × 10⁵

As can be seen in FIG. 3, there is a significant reduction in theviremia score in the group treated with the antibody. In addition, ascan be seen in FIG. 4, the percentage of infected animals in the treatedgroup are significantly lower (very low p values) as compared to theuntreated group.

-   2. Combined prophylaxis/inhibition mode—This model represents liver    transplantation. In this model mice were treated with Ab 17.1.41 (10    I.U./mouse) three days before liver transplantation followed by    transplantation of human liver fragments which were ex vivo infected    with HBV in the presence of Ab 17.1.41 (100 I.U.). HBV DNA was    tested in mice sera 10 and 17 days after transplantation. As can be    seen in FIG. 5, there was a significant reduction in the percentage    of infected animals in the treated group compared to the control    group.-   3. Combined inhibition/treatment mode—a) HBV positive human serum    was preincubated with Ab 17.1.41 followed by standard ex vivo liver    infection. b) Mice were treated with Ab 17.1.41 at days 0 and 7 post    transplantation. HBV DNA in mice sera was tested on days 11 and 18.    As can be seen in FIG. 6, the percentage of infected animals in the    Ab 17.1.41 treated group was significantly reduced but rebounded    about two weeks after the treatment was stopped.

Example 4

In the following experiment we tested the possibility to use 17.1.41 incombination with another anti viral agent in the HBV model describedabove. Mice were treated with the anti viral drug (a nucleosideanalogue, 0.5 mg/mouse/day) at days 17-20 post transplantation. A groupof mice was further treated with Ab 17.1.41 at days 19 and 20. Thepresence of HBV DNA in mice sera was tested on days 21 and 27. As can beseen in FIG. 7, immediately after treatment either with the anti viraldrug or with our monoclonal antibody there was a marked reduction in thenumber of animals infected. However, viral load rebounded in each groupthat was treated with one individual drug. Only the group that wastreated with the combination of the anti viral drug and Ab 17.1.41 didnot show an increase in the number of animals infected.

1. A human monoclonal antibody Ab17.141, which is secreted by theEuropean Collection of hybridoma cell line deposited in the CellCultures (ECACC) under Accession No. 96052169, or a fragment thereofwhich retains the antigen binding characteristics of Ab17.1.41.
 2. Thehybridoma cell line deposited at the ECACC on May 22, 1996 underAccession No.
 96052169. 3. A pharmaceutical composition for thetreatment of Hepatitis B Virus (HBV) infections comprising as an activeingredient an antibody in accordance with claim 1 together with apharmaceutically acceptable carrier.
 4. A method for the treatment ofHepatitis B Virus (HBV) infections comprising administering to anindividual in need a therapeutically effective amount of antibodiesaccording to claim
 1. 5. A method for reducing the occurrence ofHepatitis B virus (HBV) infections in a population of individuals,comprising administering a human monoclonal antibody Ab 17.1.41 or afragment thereof which retains the antigen binding characteristics of Ab17.1.41 in accordance with claim 1 to a population of individuals toreduce the occurrence of HBV infections in the population.
 6. Apharmaceutical composition for the treatment of Hepatitis B Virusinfections comprising as an active ingredient an antibody in accordancewith claim 1 adopted for use in combination with at least one otheractive ingredient being an anti viral agent.
 7. A pharmaceuticalcomposition according to claim 6 wherein the anti viral agent isselected from the group consisting of: interferons, anti-Hepatitis B(HB) monoclonal antibodies, anti HB polyclonal antibodies, nucleosideanalogues and inhibitors of DNA polymerase.
 8. A pharmaceuticalcomposition according to claim 6 wherein the anti viral agent is anucleoside analogue.
 9. A method for the treatment of HBV infectionscomprising administering to an individual in need thereof atherapeutically effective amount of a pharmaceutical compositionaccording to claim
 6. 10. A method for the treatment of HBV infectionscomprising administering to an individual in need thereof atherapeutically effective amount of a pharmaceutical compositionaccording to claim
 7. 11. A method for the treatment of HBV infectionscomprising administering to an individual in need thereof atherapeutically effective amount of a pharmaceutical compositionaccording to claim
 8. 12. A method for reducing the occurrence of HBVinfections according to claim 5, wherein the population of individualsinclude liver transplantation patients.